[Effect of VEGF165 and the VEGF aptamer pegaptanib (Macugen) on the protein composition of tight junctions in microvascular endothelial cells of the retina].
BACKGROUND
VEGF signalling is deregulated in diabetic retinopathy. Therefore, VEGF inhibitors like the modified RNA-oligonucleotide pegaptanib (VEGF aptamer, Macugen) which inhibits the interaction of VEGF(165) with its receptors, are currently being discussed as therapeutic options in the treatment of diabetic retinopathy. VEGF(165) does not only stimulate the proliferation and migration of endothelial cells but also induces delocalization of occludin which is part of the so-called tight junctions of endothelial cells likely associated with the breakdown of the blood-retina barrier.


METHODS
To further investigate the mechanisms of action of VEGF and its inhibitor, we studied the influence of VEGF(165) and/or pegaptanib on the protein composition of tight junctions in immortalised endothelial cells of the bovine retina (iBREC) by immunofluorescence staining.


RESULTS
The tight junction proteins ZO-1, occludin and claudin-5 are strongly expressed at the plasma membrane in confluent iBREC, but are located in the cytoplasm in non-confluent cells. In the presence of 50 ng/ml VEGF(165), occludin was found in the cytoplasm after 1 to 2 days, whereas claudin-5 was not and ZO-1 was only weakly influenced. However, after addition of 33 microg/ml pegaptanib for 24 h to VEGF(165)-treated iBREC, all tight junction proteins tested were again strongly expressed in the plasma membrane.


CONCLUSIONS
These results confirm an important role of tight junction proteins in the mechanisms of action of VEGF and pegaptanib on endothelial cells.
Deissler
H L
HL
Augenklinik, Universitätsklinikum Ulm. heidrun.deissler@uniklinikulm.de
Lang
G E
GE
D000818:Animals; D052157:Aptamers, Nucleotide / Q000008:administration & dosage*; D002417:Cattle; D002478:Cells, Cultured; D042783:Endothelial Cells / Q000187:drug effects / Q000378:metabolism*; D004728:Endothelium, Corneal / Q000098:blood supply* / Q000187:drug effects / Q000378:metabolism*; D008833:Microcirculation / Q000187:drug effects / Q000502:physiology*; D019108:Tight Junctions / Q000187:drug effects / Q000378:metabolism*; D042461:Vascular Endothelial Growth Factor A / Q000037:antagonists & inhibitors*
D052157:Aptamers, Nucleotide; C467484:VEGFA protein, human; D042461:Vascular Endothelial Growth Factor A; C495058:pegaptanib